Method for separating estrogen endocrine disruptors in detection environment

A technology for endocrine disruptors and estrogen, which is applied in material separation, measuring devices, and analytical materials, can solve problems such as low efficiency and heavy workload, and achieve the effect of solving the difficulty of purification and separation and improving detection efficiency

Active Publication Date: 2014-03-26
TSINGHUA UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Because there are many kinds of pollutants in sewage and only a very small part will produce estrogenic effects, it is not only a huge amount of work but also inefficient to separate and detect each pollutant one by one

Method used

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  • Method for separating estrogen endocrine disruptors in detection environment
  • Method for separating estrogen endocrine disruptors in detection environment
  • Method for separating estrogen endocrine disruptors in detection environment

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] This embodiment is a method for separating and detecting estrogen endocrine disruptors in the environment, comprising the following steps:

[0041] Step 1: Detect the binding activity of the affinity column, the specific method is as follows:

[0042] Insert 3 mL of the activated M3 strain into 20 mL of LB medium (containing 150 g / L sucrose and 50 mg / L kanamycin), and culture with shaking at 37 °C until OD 600 =0.9-1.0 (optical path=1 cm), slowly cool down to 15°C within 3 hours, add inducer 0.5mM IPTG and continue to cultivate for 16 hours, collect the bacteria by centrifugation in two parts, and freeze at -20°C. Take one of them and resuspend the bacteria with 1 mL of equilibration buffer, sonicate the bacterial cell wall (ice bath), collect the supernatant by centrifugation at low temperature, mix and incubate with the activated Ni-NTA affinity column at low temperature for 2 h, and then incubate for 3 mL of equilibration buffer to rinse. Prepare 1 mL of estradiol ...

Embodiment 2

[0049] This embodiment is a method for separating and detecting estrogen endocrine disruptors in the environment, comprising the following steps:

[0050] Step 1: Detect the binding activity of the affinity column, and calculate the recovery efficiency of the affinity column based on the recovered amount of estradiol as in Example 1.

[0051] Step 2: Separation and detection of bisphenol A by an estrogen receptor protein affinity column for the water sample containing bisphenol A standard after chlorine disinfection, the specific steps are as follows:

[0052] When the recovery efficiency of estradiol detected by the affinity column in step 1 was greater than 75%, another portion of the bacteria in step 1 was used to prepare an affinity column for estrogen receptors as described in step 1. Filter the water sample containing bisphenol A after chlorine disinfection with a 0.22 μm filter membrane, adjust its pH value to 2-3, concentrate the sample with HLB solid-phase extraction ...

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Abstract

The invention discloses a method for separating estrogen endocrine disruptors in a detection environment. The method comprises the following steps: (1) preparing a recombinant human type I estrogen receptor protein affinity chromatographic column, specifically comprising the following steps: (1-1) expressing recombinant receptor protein in genetically engineered bacteria M3, (1-2) activating a Ni-NTA affinity chromatographic column, and (1-3) collecting estrogen receptor protein; (2) separating estrogen from a mixed sample to be detected by affinity chromatography, specifically comprising the following steps: (2-1) concentrating the mixed sample, and (2-2) adding the concentrated solution into a binding buffer solution to obtain the binding buffer solution which contains the sample to be detected, wherein the premise of the amount of the added concentrated solution is no generation of precipitation; and (2-3) performing affinity separation on the estrogen endocrine disruptors from the binding buffer solution which contains the sample to be detected; and (3) performing chromatographic detection on estrogen substances in eluent.

Description

technical field [0001] The invention belongs to the technical field of analyzing toxic and harmful pollutants in water, and in particular relates to a method for separating and detecting potential estrogen endocrine disruptors in the environment. Background technique [0002] In recent years, the huge potential safety hazards caused by endocrine disruptors in the environment to public health and wild ecosystems have attracted widespread attention. Among them, estrogenic endocrine disruptors have estrogenic effects, and are the most diverse and harmful substances found so far. The largest class of endocrine disruptors. After the estrogenic endocrine disruptors in the environment enter the human body and wild animals, they will cause disorders in multiple systems including the endocrine system, reproductive system, immune system, and nervous system, resulting in reproductive disorders, individual developmental deformities, and may even cause Some species in the ecosystem beco...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/08
Inventor 陆韵商国栋李曼
Owner TSINGHUA UNIV
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